Classifications

• • C—CHEMISTRY; METALLURGY
  • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
  • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
  • C12P13/00—Preparation of nitrogen-containing organic compounds
  • C12P13/04—Alpha- or beta- amino acids
  • C12P13/08—Lysine; Diaminopimelic acid; Threonine; Valine
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23K—FODDER
  • A23K20/00—Accessory food factors for animal feeding-stuffs
  • A23K20/10—Organic substances
  • A23K20/142—Amino acids; Derivatives thereof
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23K—FODDER
  • A23K40/00—Shaping or working-up of animal feeding-stuffs
  • A23K40/10—Shaping or working-up of animal feeding-stuffs by agglomeration; by granulation, e.g. making powders
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
  • C07C229/00—Compounds containing amino and carboxyl groups bound to the same carbon skeleton
  • C07C229/02—Compounds containing amino and carboxyl groups bound to the same carbon skeleton having amino and carboxyl groups bound to acyclic carbon atoms of the same carbon skeleton
  • C07C229/04—Compounds containing amino and carboxyl groups bound to the same carbon skeleton having amino and carboxyl groups bound to acyclic carbon atoms of the same carbon skeleton the carbon skeleton being acyclic and saturated
  • C07C229/26—Compounds containing amino and carboxyl groups bound to the same carbon skeleton having amino and carboxyl groups bound to acyclic carbon atoms of the same carbon skeleton the carbon skeleton being acyclic and saturated having more than one amino group bound to the carbon skeleton, e.g. lysine

Definitions

• the invention relates to lighter and more thermally stable granular animal feed additives
• L-lysine far predominantly the L-lysine monohydrochloride is used with an L-lysine content of 78%. Since the L-lysine is prepared by fermentation, it must first be separated from all other components of the crude fermentation broth in complex process steps for the preparation of the monohydrochloride, then converted into the monohydrochloride and the latter are made to crystallize. In this case, a large number of by-products and the reagents necessary for workup fall as waste.
• EP 0 533 039 relates to processes for the preparation of an amino acid animal feed supplement
• Fermentation Brühbasis wherein the supplement can be obtained directly by spray drying from the fermentation broth.
• a part of the biomass is separated before spray drying in a variant.
• the broth can be dried even without the biomass and without additional carrier adjuvant to a manageable granules.
• L-lysine-containing solid concentrates Approximately 20 wt .-% L-lysine-containing solid concentrates are known from GB 1 439 121, in which also L-lysine-containing fermentation broths are described with a pH of 4.5 and a bisulfite content.
• EP 0 615 693 discloses a process for the preparation of an animal feed additive based on a fermentation broth, in which the fermentation broth, if appropriate after removal of part of the ingredients, is converted into a fine grain which contains at least 70% by weight maximum
• a concentrate containing L-lysine is prepared from a fermentation broth which is acidified to a pH of about 6.4 with HCl prior to concentration and to which bisulfite is added for stabilization. After evaporation, the mixture is further acidified to pH 4.0 and the desired product is obtained by spray-drying.
• EP-A 1 331 220 (US 2003/152633) relates to granulated feed additives which contain L-lysine as main component.
• the sulfate ion can be reduced by using carbon dioxide generated as a counter ion during fermentation.
• the invention relates to a granulated feed additive based on fermentation broth with
• L-lysine base 10 to 70% by weight
• L-lysine base 10 to 70% by weight
• the difference to 100 wt .-% is filled by other components of the fermentation broth and optionally the biomass.
• the granules preferably have a pH of from 3.5 to 5.1, in particular from 4.0 to 5.0, preferably from 4.2 to 4.8, measured in a 10% strength by weight suspension in deionized water at 25.degree with a pH electrode on. The measured value is constant after approx. 1 min.
• the molar ratio of sulfate to L-lysine is adjusted following the fermentation, wherein a SO 4 2- containing compounds, in particular ammonium sulfate and sulfuric acid is added in a suitable dilution.
• fertilization broth base working up an L-lysine-containing fermentation broth containing the biomass produced during the fermentation at 0 to 100%.
• the invention also provides a process for producing a granulated feed additive containing L-lysine by fermentation of an L-lysine producing microorganism in an aqueous medium using ammonium sulfate and aerobic conditions and producing a granulated product by means of actually known processes, wherein: after completion of the fermentation
• a) optionally measures the ratio sulfate / L-lysine in the fermentation broth.
• the sulfate concentration in the fermentation broth is known. In general, this also applies to the quantities of L-lysine produced under well-established conditions that the measurement is usually necessary only in case of fluctuations in production.
• the further method comprises the following steps:
• a ratio V of 1 means z.
• Step b) can also be carried out before step a).
• the L-lysine content amounts to 20 to 65, in particular 30 to 65 wt .-%.
• ammonium sulfate may then no longer be necessary. If acid is added beyond the pH reduction according to the invention, because of the buffer effect of the compounds present in the broth, increased amounts of acid are required, which can then lead to undesired denaturation and dissolution of the cells.
• the granules produced according to the invention have a pH of from 3.5 to 5.1 (measured in the suspension, see above).
• hydrochloric acid is preferably excluded. Their share generally amounts to max. 1%, in particular 0.01 to 0.1 wt .-%, based on the sulfuric acid used.
• alkali metal hydrogen sulfite preferably sodium hydrogen sulfite in an amount of 0.01 to 0.5, preferably 0.1 to 0.3, in particular 0 , 1 to 0.2 wt .-%, based on the fermentation broth to further improve.
• the sulfites, especially sodium bisulfite are preferred as a solution before the concentration of the
• Fermentation broth added.
• the amount used is preferably taken into account when adjusting the sulfate / L-lysine ratio.
• the granules are z. B. can be prepared by the method according to EP-B 0 615 693 or EP-B 0809 940, US 5,840,358 or WO 2005/006875 or WO 2004/054381. They generally have> 97% of an average particle size of> _ 0.1 to 1.8 mm and a debris weight of 600 to 950 kg / m 3 , in particular 650 to 900 kg / m 3 .
• the color values of the granules are preferably in the ranges:
• the inventive method leads not only to products with advantageous properties.
• the fermentation of the invention preferably used coryneform bacteria, in particular the genus Corynebacterium glutamicum can continuously - as described for example in PCT / EP 2004/008882 - or discontinuously in the batch process (sentence culture) or in the fed batch (feed method) or repeated fed baten method (repetitive feed process) for the purpose of producing L-lysine.
• a general summary of known cultivation methods is in the textbook by Chmiel (bioprocess 1. Introduction to bioprocess engineering (Gustav Fischer Verlag, Stuttgart, 1991)) or in the textbook by Storhas (bioreactors and peripheral facilities (Vieweg Verlag, Braunschweig / Wiesbaden, 1994) ) available.
• the culture medium or fermentation medium to be used must suitably meet the requirements suffice the respective tribe. Descriptions of culture media of various microorganisms are contained in the Manual of Methods for General Bacteriology, of the American Society for Bacteriology (Washington, DC, USA, 1981). The terms culture medium and
• Fermentation medium or medium are mutually exchangeable.
• sugars and carbohydrates such as e.g. Glucose, sucrose, lactose, fructose, maltose, molasses, sucrose-containing solutions from sugarcane or sugar cane production, starch, starch hydrolyzate and cellulose, oils and fats such as soybean oil, sunflower oil, peanut oil and coconut fat, fatty acids such as palmitic acid , Stearic and linoleic acids, alcohols such as glycerol, methanol and ethanol, and organic acids such as acetic acid. These substances can be used individually or as a mixture.
• sugars and carbohydrates such as e.g. Glucose, sucrose, lactose, fructose, maltose, molasses, sucrose-containing solutions from sugarcane or sugar cane production, starch, starch hydrolyzate and cellulose, oils and fats such as soybean oil, sunflower oil, peanut oil and coconut fat, fatty acids such as palmitic acid , Stearic and lin
• organic nitrogen-containing compounds such as peptones, yeast extract, meat extract,
• the nitrogen sources can be used singly or as a mixture.
• phosphorus source can phosphoric acid
• the culture medium must further contain salts, for example in the form of sulphates of metals such as, for example, sodium, potassium, magnesium, calcium and iron, for example magnesium sulphate or iron sulphate, which are necessary for growth.
• salts for example in the form of sulphates of metals such as, for example, sodium, potassium, magnesium, calcium and iron, for example magnesium sulphate or iron sulphate, which are necessary for growth.
• essential growth factors such as amino acids, for example, homoserine and Vitamins such as thiamine, biotin or pantothenic acid be used in addition to the above substances.
• suitable precursors of the respective amino acid can be added to the culture medium.
• the said starting materials can be added to the culture in the form of a one-time batch or fed in a suitable manner during the cultivation.
• the fermentation at elevated pressure, for example at a pressure of 0.03 to 0.2 MPa, driven.
• the temperature of the culture is usually 20 ° C to 45 ° C, and preferably 25 ° C to 40 ° C.
• the cultivation is continued until a maximum of the desired amino acid has formed. This goal is usually reached within 10 hours to 160 hours. In continuous processes longer cultivation times are possible.
• the invention accordingly provides a process for preparing an L-amino acid in which
• the L-amino acid accumulates in the fermentation broth or in the cells of the isolated coryneform bacterium.
• the fermentation broth thus prepared is then further processed into a solid or liquid product.
• a fermentation broth is understood as meaning a fermentation medium in which a microorganism has been cultivated for a certain time and at a certain temperature.
• the fermentation medium or the medium used during the fermentation contains / contain all substances or components which ensure an increase of the microorganism and a formation of the desired amino acid.
• the resulting fermentation broth contains accordingly a) the biomass of the microorganism resulting from the multiplication of the cells of the microorganism, b) that during the course of the fermentation
• the organic by-products include substances which are optionally produced by the microorganisms used in the fermentation in addition to L-lysine and optionally excreted. Pay for it L-amino acids, which account for less than 30%, 20% or 10% compared to the desired amino acid. These include organic acids which carry one to three carboxyl groups such as
• acetic acid for example, acetic acid, lactic acid, citric acid, malic acid or fumaric acid.
• sugar such as trehalose.
• Typical fermentation broths suitable for industrial use have an L-lysine content of 40 g / kg to 180 g / kg or 50 g / kg to 150 g / kg.
• the content of biomass (as dried biomass) is generally 20 to 50 g / kg.
• the biomass may be wholly or partially separated by separation methods such as e.g. centrifugation, filtration, decantation, or a combination thereof, from the fermentation broth or left entirely in it.
• separation methods such as e.g. centrifugation, filtration, decantation, or a combination thereof, from the fermentation broth or left entirely in it.
• the biomass or the biomass-containing fermentation broth is inactivated during a suitable process step.
• the biomass is completely or almost completely removed so that no (0%) or at most 30%, at most 20%, at most 10%, at most 5%, at most 1% or at most 0.1% biomass remains in the manufactured product ,
• the biomass is not or only slightly removed so that all (100%) or more than 70%, 80%, 90%, 95%, 99% or 99.9% biomass remains in the product produced.
• Fermentation broths from which the biomass has been partially or totally removed can be used to standardize the product. This naturally also applies to the pure compounds L-lysine base and lysine sulfate.
• the fermentation broth obtained after the fermentation is acidified with sulfuric acid before the concentration and optionally mixed with ammonium sulfate.
• the broth can also be stabilized and lightened by the addition of preferably sodium hydrogen sulfite or another salt, for example ammonium, alkali metal or alkaline earth metal salt of the sulfurous acid.
• biomass is separated off, this is preferably carried out before the inventive lowering of the pH and the addition of ammonium sulfate and sulfite salt.
• Biomass are optionally partially or completely removed in the fermentation broth contained organic or inorganic solids.
• (Starting materials) remain at least partially (> 0%), preferably at least 25%, more preferably at least 50% and most preferably at least 75% in the product. If appropriate, these also remain completely (100%) or almost completely ie> 95% or> 98% in the product.
• the term "fermentation broth base" means that a product contains at least a portion of the components of the fermentation broth. Subsequently, the broth is removed or thickened or concentrated by known methods, such as, for example, with the aid of a rotary evaporator, thin film evaporator, falling film evaporator, by reverse osmosis or by nanofiltration.
• This concentrated fermentation broth can then be worked up by freeze-drying, spray-drying, spray granulation or by other methods, for example in the circulating fluidized bed according to PCT / EP 2004/006655, into free-flowing products, in particular granules.
• a product having the desired grain size is isolated from the resulting granules by sieving or dust separation.
• Particle size determination can be performed by laser diffraction spectrometry techniques.
• the corresponding methods are in the textbook for
• the free-flowing, finely divided powder can in turn be converted by suitable compacting or granulating process into a coarse-grained, readily pourable, storable and substantially dust-free product.
• Free-flowing refers to powders that flow out of a series of glass outlet vessels with different sized outflow openings at least from the vessel with the opening 5 mm (millimeters) unhindered (Klein: Soaps, Ole, Fats, Wachse 94, 12 (1968)).
• finely divided is meant a powder having a predominant proportion (> 50%) of a particle size of 20 to 200 ⁇ m in diameter.
• coarse-grained is meant a product having a predominant proportion (> 50%) of a particle size of 200 to 2000 ⁇ m in diameter.
• dust-free means that the product contains only small amounts ( ⁇ 5%) of particle sizes below 100 microns in diameter.
• auxiliaries such as starch, gelatin, cellulose derivatives or similar substances, such as are commonly used in food or feed processing as binders, gelling or thickening agents, or of other substances such as silicas, silicates (EP-A 0 743 016) stearates.
• oils mineral oils vegetable oils or mixtures of vegetable oils can be used. Examples of such oils are soybean oil, olive oil, soybean oil / lecithin mixtures.
• silicone oils, polyethylene glycols or hydroxyethycellulose are also suitable.
• the proportion of dust, ie particles with a particle size ⁇ 100 microns is preferably> 0 to 1 wt .-%, more preferably at most 0.5 wt .-%.
• the product can also be applied to a known and customary in the feed processing organic or inorganic carrier material such as silicas, silicates, shot, brans, flours, strong sugar or other and / or mixed with conventional thickening or binding agents and stabilized.
• organic or inorganic carrier material such as silicas, silicates, shot, brans, flours, strong sugar or other and / or mixed with conventional thickening or binding agents and stabilized.
• the product can also be obtained by coating processes with film formers, such as, for example, metal carbonates, silicas, silicates,
• the corresponding amino acid can be added during the process in the form of a concentrate or, if appropriate, a substantially pure substance or its salt in liquid or solid form. These can be added individually or as mixtures to the obtained or concentrated fermentation broth, or also during the drying or granulation process.
• the fermentation broth-based solid product thus prepared has a lysine content (calculated as lysine base) of 10% to 70% by weight, preferably 30% to 60% by weight, and most preferably from 40% to 60% by weight based on the total amount of the product.
• a lysine content calculated as lysine base
• the adjustment of the pH value in the fermentation broth to values ⁇ pH 5.2, the increase of the sulphate / lysine ratio and preferably a sulphite addition in the range from 0.01 to 0.5 wt .-% in the fermentation broth after fermentation significantly reduces lysine loss during processing of the fermentation broth.
• concentration of the concentrate results in an average lysine loss of about 3.5% by weight (without granulation step).
• the increase in the sulfate ratio by adding ammonium sulfate leads in the end to an average loss of lysine of about 3.2% by weight, and the sole adjustment of the pH reduces the average loss of lysine to about 1.4% by weight.
• the combination of pH adjustment and increasing the sulfate ratio shows a higher protective effect for the lysine and results in an average lysine loss of about 0.9 wt .-%.
• the combination of pH adjustment and Natriumhydrogensulfitzugabe, together with the combination of all three additives only an average lysine loss of about 0.6 wt .-% or about 0.7 wt .-%.
• the 3-range colorimeter for color and remission measurement works according to the principle described in DIN 5033, whereby the diffuse reflection of the sample is measured at an angle of 8 °.
• the reflected light is transmitted via a light guide into the device for splitting onto the precisely defined standard color filter. Measured against a calibration standard.
• Table 1 shows the results of the color determination for the comparative experiments A to D. Even without acidification of the fermentation broth, products having an acidic pH are obtained, but their color values do not reach those of the products according to the invention.
• Samples E and F correspond to the products according to the invention, which were obtained after acidification of the broth to pH 5.1, wherein the broth used to prepare sample F additionally 0.2 wt .-% sodium bisulfite were added.
• Table 2 shows the superiority of the inventive products E and F under thermal stress with regard to a lower L-lysine loss.
• E weight of sample in g Note: If you want to monitor the water absorption over a longer period, a water intake curve is determined. For this purpose, the weight of the weathered sample is in the first 6 hours hourly and once after 24 hours.
• Figure 1 shows that the inventive product E absorbs lower amounts of water / time than the products A to D according to the prior art.
• Table 2 Stability under thermal stress (85 ° C).
• Granulation was carried out according to EP-B 0 809 940 (US 5,840,358).
• the concentrated broth was granulated according to EP-B 0 809 940.
• the pH was lowered by the addition of 0.54 kg sulfuric acid (about 93%) to pH 5.2, so that the initial L-lysine content of 57.7% to 55.7 wt .-% decreases.
• the resulting fermentation broth was heated to 55 ° C and then concentrated as in Example 2.1 and granulated.
• the sulphate / L-lysine ratio after fermentation was 0.95, so that no further ammonium sulphate was metered in any more.
• the resulting fermentation broth was heated to 55 ° C. and then concentrated and granulated as in Example 2.1.

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  • 2006-04-06 DE DE102006016158A patent/DE102006016158A1/de not_active Withdrawn
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